SusChEM: Collaborative proposal: Engineering increased activity of cutinase toward poly(ethyleneterephthalate) for recycling of plastic

SusChEM：合作提案：通过工程设计提高聚对苯二甲酸乙二醇酯的角质酶活性，以回收塑料

• 批准号: 1930825
• 负责人: Romas Kazlauskas
• 金额: $ 9.27万
• 依托单位: University of Minnesota-Twin Cities
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2023-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1930825&HistoricalAwards=false
• 关键词: SusChEM; Collaborative; proposal; Engineering; increased

The environmental consequences of plastic solid waste are serious. The problem continues to increase as we diversify and expand plastic use and applications. The most significant example of this problem is the rate that plastics are accumulating in oceans, lakes and rivers. The most widely used plastic is polyethylene terephthalate, or PET. PET is used in the manufacture of bottles, polyester fabrics, and food trays. Production levels are estimated to be over 50 million tons per year. This project will attempt to develop an efficient enzyme for the degradation of PET. High school students will be engaged in summer research opportunities. Undergraduate and graduate students will also be involved in the project. These opportunities will help develop a STEM workforce.Recently, several groups discovered enzymes that catalyze PET hydrolysis under mild conditions. Unfortunately, these enzymes are too slow and too unstable for practical use. Protein engineering to increase their catalytic efficiency is a potential solution. The project tests a new approach for engineering enzymes to act on insoluble substrates. This approach recognizes that the interaction between an enzyme and an insoluble substrate is complex and involves an extended binding site. The enzyme must pull the chain from the bulk polymer, position the ester group for hydrolysis, and allow the polymer substrate to slide within the binding site for repeated hydrolysis of the chain. Previous attempts to increase the activity of enzymes toward insoluble substrates have met with limited success. The proposed large-area-mutagenesis (LAM) methodology to be implemented in this program is a systematic strategy for engineering enzymes for insoluble synthetic polymer substrates. The approach will: 1) establish the size of the binding region in cutinase that influences its catalytic activity toward PET; 2) optimize the shape of the extended binding region for PET, 3) increase the thermal stability of the enzyme for use above the glass transition temperature of PET, where chain mobility is higher, allowing better access of the enzyme to the polymer substrate. The results of this program will be the engineering of an efficient, stable enzyme with increased catalytic activity of at least 100-fold that may be suitable for commercial PET recycling, which is an essential step to reducing plastic pollution. This project is supported jointly by the Cellular and Biochemical Engineering Program in CBET/ENG and the Chemistry of Life Processes Program in CHE/MPS.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

塑料固体废物的环境后果是严重的。随着我们多样化并扩大塑料使用和应用，问题继续增加。这个问题的最重要的例子是塑料在海洋，湖泊和河流中积累的速度。使用最广泛的塑料是聚对苯二甲酸酯或宠物。 PET用于制造瓶子，聚酯织物和食物托盘。估计生产水平每年超过5000万吨。该项目将尝试开发一种有效的酶来降解宠物。高中生将参与夏季研究机会。本科生和研究生也将参与该项目。这些机会将有助于发展STEM劳动力。几个小组发现在轻度条件下催化PET水解的酶。不幸的是，这些酶太慢且不稳定，无法实际使用。蛋白质工程以提高其催化效率是一种潜在的解决方案。 该项目测试了一种新的工程酶对不溶性底物作用的方法。这种方法认识到酶与不溶性底物之间的相互作用是复杂的，并且涉及扩展的结合位点。酶必须从大量聚合物中拉出链，将酯基团放置进行水解，并允许聚合物底物在结合位点中滑动以重复链的水解。以前的尝试增加了酶对不溶性底物的活性的尝试有限。该程序中要实施的拟议的大面积 - 助剂（LAM）方法是一种系统的策略，用于工程酶的不溶性合成聚合物底物。方法将：1）建立切碎酶中结合区域的大小，从而影响其对PET的催化活性； 2）优化PET的扩展结合区域的形状，3）增加酶的热稳定性，用于在PET的玻璃过渡温度之上使用，其中链迁移率更高，从而使酶更好地进入聚合物底物。该程序的结果将是一种高效，稳定的酶的工程，其催化活性至少为100倍，可能适合商业宠物回收，这是减少塑料污染的重要步骤。该项目得到了CBET/ENG的蜂窝和生化工程计划的共同支持，以及CHE/MPS中的化学过程计划计划，该奖项反映了NSF的法定任务，并被认为值得通过基金会的知识分子优点和更广泛的影响审查标准通过评估来获得支持。